This application seeks to study a subpopulation of activated macrophages that we've identified with potent immunoregulatory activity. We will determine if we can exploit these regulatory macrophages (R-Mf) to develop a novel class of anti-inflammatory therapeutics. In Aim 1, we will characterize regulatory macrophages, and identify a panel of R-Mf-specific biomarkers to develop a signature for these cells. These biomarkers will be used to determine the various reprogramming signals that can give rise to regulatory macrophages and they will enable the identification of these cells in tissue during disease. We propose that the depletion or induction of R-Mf represent new approaches to treat diseases. In Aim 2, we will examine the conversion of classically activated macrophages (Ca- Mf) to regulatory macrophages. We propose that Ca-Mf can control their own activation state by secreting ATP and rapidly converting it to adenosine. Adenosine induces a regulatory macrophage phenotype. In the absence of this conversion, macrophage activation progresses uncontrolled, leading to inflammatory pathology. This represents a new paradigm in (controlling) macrophage activation. We will determine the role that macrophage ectoenzymes, CD39 and CD73, play in promoting the conversion of ATP to adenosine by engineering macrophages to overexpress or lack these ectoenzymes. The biomarkers developed in Aim 1 will help us to identify the induction of regulatory macrophages following their exposure to adenosine. In Aim 3, we will manipulate macrophages to improve human health. We will induce the formation of R-Mf and determine whether the reversal of disease pathology correlates with the induction of regulatory macrophages. We will utilize the biomarkers developed in Aim 1 to identify the persistence of R-Mf in tissue during the process of disease resolution. We will generate R-Mf by either adding exogenous 'reprogramming' signals (Aim 1) or by promoting the conversion of ATP to adenosine (Aim 2). The core hypothesis to be tested in these studies is that macrophage physiology can be reliably and predictably manipulated, and that R-Mf can be exploited to modify immune responses and affect disease outcomes. By inducing regulatory macrophages we can prevent or reverse autoimmune pathologies. By deleting regulatory macrophages we may enhance immunity.